Abstract—Protein microarrays represent a promising alternative in proteomics and in the biomedical industry. The widespread use of this technology has been limited, largely due to the labor intensive protein production, the quality of proteins expressed in different systems, and the shelf life of these arrays. The novel method Nucleic Acid Programmable Protein Array (NAPPA) overcomes these limitations by synthesizing the proteins in situ. NAPPA entails spotting plasmid DNA encoding the relevant proteins, which are then simultaneously transcribed and translated by a cell free system. NAPPA assays are based on fluorescence detection, which need a protein labeled, often in-situ or by fluorescent tag, and can’t give real time information or kinetics. An attractive alternative to traditional fluorescence-based microarrays detection methods is the technique of surface plasmon resonance imaging (SPRi). SPRi brings a significant advantage in the analysis of biological samples where labeling multiple biomarker with fluorophores or nanoparticles is not practical. A new surface chemistry is developed to carry out an in-situ and cell free synthesis of an SPR compatible protein array. The DNA which encodes the P53-GST-(e-coil) fusion protein is arrayed on the gold sensor surface and through the expression with cell lysate extract the corresponding protein array is obtained. Both the expression process and the posterior characterization are implemented on the flow cell covering the sensor chip. The kinetic interaction of the fusion proteins with the specific antibody anti-P53 was analyzed.
Index Terms—Cell free, in-situ, protein array, SPRi
Javier Batista Pérez, Amit Nand, and Zhu Jinsong are with the National Center for Nano science and Technology, China, Beijing 100080, China
Javier Batista Pérez is with the Centro de Estudios Avanzados, Habana 19370, Cuba (e-mail: jbatista08@gmail.com)
Cite: Javier Batista Pérez, Amita Nand, and Zhu Jinsong, "Integrated Method of in Situ Cell Free Synthesis of a Protein Array on Gold Surface and Real Time Kinetic Monitoring by SPR Imaging," International Journal of Engineering and Technology vol. 4, no. 5, pp. 500-503, 2012.
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